‘Single-shot’ pulmonary vein isolation using a novel lotos pulsed field ablation catheter: a pre-clinical evaluation of feasibility, safety, and 30-day efficacy

Abstract Aims Pulsed field ablation (PFA) is emerging as a non-thermal, tissue-specific technique for pulmonary vein isolation (PVI) in atrial fibrillation therapy. This pre-clinical study aims to investigate the feasibility and safety of PVI using a novel PFA system including a nanosecond-scale PFA generator, a novel lotos PFA catheter, and a customized 12 Fr steerable sheath. Methods and results A total of 11 Yorkshire swine were included in this study, with 4 in the acute cohort and 7 in the chronic cohort. Under general anaesthesia, transseptal puncture and pulmonary vein (PV) angiography was initially performed. The PFA catheter was navigated to position at the right and left PV antrum after the electroanatomic reconstruction of the left atrium. Biphasic PFA applications were performed on PVs in both the spindle-shaped and the lotos-shaped poses. Pulmonary vein isolation and PFA-associated safety were assessed 30 min after ablation in both cohorts and 30 days later in the chronic cohort. Detailed necropsy and histopathology were performed. Additional intracardiac echocardiography and coronary angiogram were evaluated for safety. All target PVs (n = 20) were successfully isolated on the first attempt. No spasm of coronary artery or microbubble was seen during the procedure. Eleven of 12 PVs (91.6%) remained in isolation at the 30-day invasive study. No evidence of PV stenosis was observed in any targets. However, transient diaphragm capture occurred in 17.6%. Histopathological examinations showed no evidence of collateral injury. Conclusion This study provides scientific evidence demonstrating the safety and efficacy of the novel PFA catheter and system for single-shot PVI, which shows great potential.


Introduction
2][3][4] However, the currently used classic ablation techniques, radiofrequency (RF) and cryoballoon (CB) ablation, cannot meet the clinical demands of cardiomyocyte-specific damage. 5on-selective damage of vascular smooth muscle cells has long been a shadow of thermal energy ablation, making pulmonary vein (PV) stenosis a serious complication following traditional cardiac ablation. 6,7dditionally, adjacent anatomical structures such as the ganglionated plexus or the oesophagus may be affected during the procedure, resulting in abnormal cardiac autonomic regulation and/or oesophageal injury. 5,8ulsed field ablation (PFA), with its safety and efficacy demonstrated in numerous publications, [9][10][11][12][13][14] has shown PFA as a potential alternative in the future.6][17] Moreover, it has been proved that approaches using 'single-shot' circumferential PVI (CPVI) are more attractive than point-by-point approaches due to their simpler workflow and shorter procedure time. 5,18Thus, a single-shot PFA catheter holds great promise for AF therapy, given its feasibility for CPVI and its specific feature of causing cardiomyocyte-selective damage.It is recognized, however, that the operating convenience and the energy control of current PFA catheters are quite inadequate.][21][22] Furthermore, different catheter designs and pulse parameter settings can impact the catheter's contact with tissue and the number of recommended applications (e.g.whether the rotation is required for additional applications).It also plays an essential role in interfering with collateral tissues (e.g.capturing the phrenic nerve). 19,23Consequently, outcomes in previous assessments of current PFA catheters are not duplicable in terms of effectiveness and safety for new products.
In this pre-clinical study, we evaluated the feasibility and safety of a novel PFA system.The system includes a nanosecond-scale pulsed field generator, an 11 Fr over-the-wire expandable lotos catheter for singleshot PVI, and a customized 12 Fr steerable sheath.The catheter design and the nanosecond-scale energy delivery parameters simplify the manipulation and minimize the risk of capturing the phrenic nerve during the procedure.The effectiveness of the system was evaluated by measuring PV potentials, PV bidirectional blocking, and doing electroanatomic mapping before, immediately after, and 30 days after ablation.Gross pathology, histopathology, as well as surrounding tissue injury, are assessed simultaneously.

Animals and protocols
All the experiments were performed in Guangdong Jinshi Medical Technology Service Co., Ltd and were approved by the Institutional Animal Care and Use Committee (IACUC) of the Guangdong Jinshi Medical Technology Service Co., Ltd (approval number IACUC-T-2022-013).All procedures followed the Guideline for the Care and Use of Laboratory Animals of Guangdong Provincial People's Hospital.A total of 11 Yorkshire swine were included in this study, with 4 in the acute cohort and 7 in the chronic cohort.

Pulsed field ablation system and catheter
The ablation system in this study comprised a PFA generator (InRythm, Insight Lifetech, Shenzhen, China), a proprietary lotos-shaped PFA Catheter (LotosPFA, Insight Lifetech), and a customized 12 Fr steerable sheath (InBridge, Insight Lifetech) to navigate and position the PFA catheter.The PFA generator is capable of delivering high-voltage pulsed field waveforms over multiple channels with variable amplitudes.The 11 Fr over-the-wire PFA ablation catheter with 16 electrodes was arranged in a lotos configuration when being fully deployed, with a transition state as a spindle (Figure 1A1-A3).As shown in Figure 1B, each electrode was assigned a specific number.Biphasic waves are generated by electrode pairs consisting of one odd-numbered electrode and one even-numbered electrode.All 16 electrodes were capable of recording bipolar electrograms, as well as pacing.The catheter tip was advanced over a 0.035 J-tipped guidewire to reach the PV ostia in its spindle shape and achieve optimal contact with the PV antrum with its lotos framework.The catheter was available in two sizes, with tip diameters of 28 and 31 mm in lotos pose, respectively.This allows for positioning the catheter tip at the PV antrum effectively and achieving appropriate contact, regardless of anatomic variations.
Trains of nanosecond-scale pulses were delivered in a bipolar, biphasic fashion in each electrode pair (Figure 1D).All electrode pairs were employed in sequence during one delivery process.Arcing and muscular motion were avoided during the delivery of PFA by using a proprietary optimized catheter and programme design including electrode size, spacing, pulse width, and voltage amplitude.
Pulsed field ablation waveforms consist of a sequence of nanosecond-scale pulses, with an amplitude of 1850 V for the spindle pose and 2100 V for the lotos pose.The delivery procedure for each vein contains two to four applications in the spindle pose and additional two to four applications in the lotos pose, which depends on the operator's judgement.The total application duration is <1 min for one target.

Procedural details
The swine ablation model was described previously. 19In brief, 4-month-old Yorkshire swine (50-60 kg) were administered general anaesthesia with mechanical ventilation.Rivaroxaban (150 mg orally every day) was administered for 7 days before PFA.A constant irrigation of the PFA catheter was maintained throughout the procedure to prevent thrombosis.The procedures were conducted under heparin anticoagulation with an activated clotting time range of 300-350 s.
A coronary sinus multi-electrode catheter was placed via left femoral access for sensing and pacing.A single transeptal puncture was performed with the 12 Fr custom-steerable sheath (InBridge, Insight Lifetech) for left atrial access by using a standard transeptal technique.Pulmonary venous angiography was performed to obtain baseline PV diameter.HD Grid (Abbott, Chicago, IL, USA) was used for activation and voltage mapping of the left atrium (LA) with the EnSite™ NavX™ Precision Three-dimensional mapping system (Abbott).
The PFA catheter was navigated into the LA and the wire tip was positioned within a distal PV branch.The catheter tip was advanced into the right superior PVs (RSPVs) and left superior PVs (LSPVs) and then expanded gradually by operating the actuator in the handle under fluoroscopic guidance.Tissue contact was judged only by resistance during catheter push manoeuvres and trains of nanosecond-scale biphasic pulses were delivered in the spindle shape in the PV ostia and then in the lotos shape in the PV antra.There is no requirement for these pulses to be synchronized with either atrial or ventricular depolarization.
For immediate and 30-day efficacy assessment, high-density voltage maps (including the assessment of vein potentials) were prepared using the HD Grid catheter after the 30 min waiting period and 30 days later, along with a PV bidirectional block test.
For safety assessment, venous angiography was performed to assess the diameter of each vein before and after ablation.In addition, intracardiac echocardiography (ICE) was used to inspect the presence of microbubbles generated during ablation.A coronary angiogram was performed simultaneously with ablation in several swine in order to evaluate the impact of PFA on coronary arteries.Once all assessments were completed, the study catheter was removed and examined for thrombus or tissue attachment, and ICE was conducted to inspect the presence of iatrogenic pericardial effusion.

Follow-up and histological investigation
All acute cohort swine were humanely sacrificed on the same day after completing all procedures.All swine in the chronic cohort were recovered and monitored for 30 days.A repeat procedure including electroanatomic mapping, PV potential recording, PV bidirectional block test, and PV angiography was conducted after 30 days.Both cohorts of swine were euthanized, and their hearts and adjacent tissues were collected for gross pathological and histopathological analyses at the end of their predetermined survival periods.
Upon documentation of any abnormalities or injuries, all explanted hearts and neighbouring organs were fixed in formalin.The ablated PVs were identified, opened along their long axes, and trimmed evenly along the vein axis to obtain four circumferential longitudinal sections per PV.Samples were embedded in paraffin, sectioned into slides, and stained with Haematoxylin and Eosin and Masson's Trichrome.
A veterinary pathologist blinded to the electroanatomic data and outcomes assessed the slides.The sections of veins with myocardial sleeves were carefully assessed for the presence and transmurality of lesions, extent of fibrosis, damage to nerves, arterioles, and venules within the lesion, presence of oedema, level of inflammatory response, presence of thrombi or haemorrhage, and any other relevant findings.
The oesophagus and other adjacent tissues were carefully examined for gross pathology and photographed.Since no abnormalities were visible on the surface, no area of interest was sectioned for histological analysis.

Statistical analyses
Continuous variables were presented as mean ± standard deviation (SD), whereas categorical variables were presented as count and percentage.A two-sided Student's t-test was employed to compare continuous variables between two groups.In the case of comparisons among more than two groups, one-way analysis of variance (ANOVA) analysis and the Tukey multiple comparisons test were used, as appropriate.Categorical variables were compared using χ 2 analysis or Fisher's exact test, as appropriate.A P value <0.05 was considered statistically significant.Statistical analyses were performed with SPSS 24.0 software (SPSS Inc., Chicago, IL, USA), and graphs were generated using GraphPad Prism software (version 9.1.1).

Procedural outcomes
The study catheter was successfully navigated into all targets with the spindle pose at the PV ostia and the lotos pose at the PV antrum, as confirmed by the fluoroscope (Figure 2).Pulsed field ablation was delivered to all targeted PVs, with an average of 3.7 ± 0.8 applications in the spindle pose and 3.5 ± 0.9 applications in the lotos pose.The number of applications for each PV is shown in Supplementary material online, Table S1.Post-ablation and 30-day follow-up assessments were successfully performed.

Acute cohorts
In four swine, all targeted PVs (four RSPVs and four LSPVs) were successfully isolated at the first attempt (Table 1, Figure 3A-C).Electroanatomic maps showed low-voltage areas (<0.1 mV) when the target PVs were remapped after a 30 min waiting period.In general, acute PVI was achieved in 100% of PVs.

Chronic cohorts
The LSPVs of two swine were excluded due to the difficulties of catheter assessment because of the low-voltage areas of the PV antra shown in baseline electroanatomic maps.Similar to acute cohorts, all targeted PVs, including seven RSPVs and five LSPVs, achieved acute isolation, which was assessed by PV potential recording, bidirectional block test, and electroanatomic mapping at baseline and after the 30 min waiting period (Figure 3A-E).All swine completed a follow-up period averaging 32.1 ± 2.0 days.In the 30-day follow-up, reassessment revealed short-term isolation in five out of five LSPVs (100%) and six out of seven RSPVs (85.7%;Table 1).Overall, the 30-day efficacy rate of PVI in this cohort was 91.6%.

Safety assessment
No catheter placement or deployment-associated complications occurred during the procedure (e.g.pericardial effusion, cardiac tamponade, phrenic palsy, or evidence of an air embolism).No significant difference in PV diameter between the pre-ablation and 30 min postablation groups was observed.Similar results were found in the 30-day follow-up group compared with the pre-ablation group (Table 2, Figure 4A and B).
During PFA deliveries, neither phrenic palsy, notable arrhythmias, nor ST-segment elevations occurred.The coronary angiography was performed concurrently with PFA in two swine, and no evidence of coronary spasm was observed (Figure 4C).No microbubbles were detected by ICE during PFA (Figure 4D, see Supplementary material online, Video S1).Neither symptoms of phrenic palsy nor cerebral embolism were detected during the follow-up period.
However, transient phrenic nerve capture was observed in several cases, especially when the PFA catheter was expanded in the spindle pose (Table 3).In general, phrenic nerve capture occurred in 29.4% of PFA applications in the spindle pose, and 5.9% in the lotos pose, resulting in a total of 17.6%.Phrenic nerve capture was more frequently observed in the LSPV than in the RSPV (27.3 vs. 8.6%).

Histological investigation
The entire hearts and adjacent tissues of all swine were successfully extracted, showing macroscopic lesion formation at each target area in each animal.On the endocardial surface, lesions were identified as white patches (Figure 5A).No ablation-related macroscopic lesions were observed in neighbouring organs, including phrenic nerves and oesophagus (see Supplementary material online, Figure S1).
Because of the limited scope of gross pathology, a histological examination was conducted.As shown in Table 1, histological analysis of all sections of LSPVs and RSPVs confirmed the presence of ablation lesions (Figure 5B-D).However, in the chronic cohort, 47 out of 48 (97.9%) sections achieved transmural lesions.The mean lesion thickness was 0.82 ± 0.54 mm in LSPVs and 1.32 ± 0.60 mm in RSPVs.Only one LSPV did not achieve transmural lesion.
All lesions caused by PFA were continuous, homogenous, and endocardial sparing, with the structural framework preserved in all samples (Figure 5B-F).The myocardium was replaced by fibrosis within the lesion.Neither the arteries, veins, nor nerves within the lesion were damaged (Figure 5C and E).Endocardial oedema was present in acute histopathology (Figure 5F) but disappeared in histological sections after 30 days.Haemorrhage and myocardium inflammation were observed in several cases (Figure 5B).
No serious IRE-induced complications, such as thrombus, were observed in any of the samples.All the results of the histopathology are summarized in Table 1.

Discussion
In this study, we assessed the feasibility and safety of a lotos PFA catheter as well as a proprietary PFA system designed to achieve 'singleshot' PVI for AF.The main findings can be summed up as follows.(i) The catheter was successfully delivered to and contacted all target veins.The procedural workflows were simple and time-saving with a short learning curve for operators.(ii) Pulmonary vein isolation was achieved with a 100% instant success rate in both cohorts.The 30-day isolation rate was 91.6%, indicating excellent effectiveness of the study catheter as well as the PFA system.(iii) No instances of PV stenosis, oesophagus injury, coronary spasm, or microbubbles were observed during and after the procedures, suggesting a level of acceptable safety of the study catheter.(iv) Phrenic nerve capture was observed in 17.6% of applications during ablation with no phrenic palsy in the follow-up period, indicating transient capture rather than sustained injury.(v) In histopathology, the lesions were transmural, continuous, homogenous, and collateral tissue-sparing, with the preservation of the structural framework of the heart.Together, these findings demonstrate that the study catheter and the PFA system are effective and safe for PVI, showing great potential for future clinical applications.

Catheter design and workflow
This catheter configuration and nanosecond electric pulse setting play a significant role in the effectiveness and safety of ablation.According to previous literature, energy delivery depends on a variety of energy parameters, including pulsed amplitude (voltage), pulse duration, number of pulses, and pulse repetition rate. 17,23,24It has been reported that efficacy and tissue damage are dose-dependent, 25,26 highlighting the importance of carefully tailoring PFA parameters to avoid collateral damage.We acknowledge that the study catheter and the workflow resemble the ones from flower-shaped PFA catheters including Farawave (Farapulse Inc., CA, USA), but there is indeed quite a difference in terms of catheter structure, parameter settings, energy delivery, and manoeuvering procedure.(i) Although the fully deployed catheter tips are both flower-liked, this study catheter owns more splines and each of the two adjacent splines is linked with electrodes.The novel design contributes to a more stable structure, making the catheter reach the targets easily even with sharp turns as well as enabling optimal contact with various PV ostia diameters.(ii) We also employed a biphasic wave, as in other studies, [19][20][21][22]27,28 to achieve effective treatment with a reduced risk of adverse reactions (e.g. an unwanted temperature increase, phrenic nerve stimulation). (iii)The pulses we generate are on a nanosecond scale.This delivery pattern shortens the delivery time and moderates intracellular effects.[29][30][31] In addition, although the underlying explanation is unclear, it reduces the likelihood and extent of phrenic nerve stimulation, as we have observed (see Supplementary material online, Video S2).(iv) In terms of operation, the novel lotos design eliminates the need for rotation during PFA deliveries, simplifying the workflow.
Other specificities include the material of the catheter tip, which is made of a flexible material that allows for small catheter deformation that can better fit the PVs in real-world applications.Additionally, there are safety apparatuses.Prior to advance into the LA, an impedance and safety test is performed, and a forced fusion mechanism is available to prevent unintended ablations in time.
A single application for each target has been deemed ideal for PFA innovation.Despite this, there are currently no reliable data on the long-term efficacy of PVI with only one application of PFA energy delivery.Recently, Koruth et al. reported that isolation was achieved in 2/2 RSPVs but not in SVCs with one application after a 1 week follow-up period.According to our previous small-sample pilot study, 6/6 PVs achieved isolation after 19 days, while 3/4 PVs exhibited durable isolation after 84 days (see Supplementary material online, Table S2).However, damage to the PV antrum was relatively small in the long-term remap (see Supplementary material online, Figure S2), suggesting that a single application may not meet the clinical requirements.Therefore, the current protocol was chosen for this study.Further studies are required in order to optimize the catheter and parameters.3][34] Howard et al. 25 conducted biphasic, bipolar PFA on an isolated porcine heart model and found that the size of the lesions decreased as the distance between the epicardial surface and the electrodes increased. 26,32It is important to note that lesions can form in the absence of direct electrode contact with tissue, making PF delivery more flexible than RF or cryoablation.However, a lack of electrodetissue contact can result in inadequate lesion formation and ineffective ablation. 35Interestingly, basic experiments suggest that achieving tissue contact is more important than the force of contact. 34Our study indicates that complete contact is required in order to achieve transmural damage.
The updated literature reports that the 34-mm floral catheter tip (FARAWAVE™, Boston Scientific) increased the incidence of postoperative arrhythmias.Electroanatomic mapping suggested macroreentry atrial tachyarrhythmia with a critical isthmus at the left atrial posterior wall.In the present study, the catheter tips are available in two sizes, 28 and 31 mm.Neither post-operative arrhythmias nor reconnection of the left atrial posterior wall on electroanatomic mapping were recorded during the follow-up period.However, for the purpose preventing unintentional roof line block, further research is necessary in order to identify the optimal range of tip sizes for clinical use.
Overall, the proprietary catheter design and optimized pulse parameters allowing 'single-shot' PVI meet the requirements of safety and 30-day efficacy.

Phrenic nerve capture
In our study, we observed several cases of phrenic nerve capture during pulsed electric field discharges (Table 3).This event was associated with the catheter position and the pulse parameters, based on our observation.A previous study has shown that the response of the phrenic nerves is dose-dependent and is affected by the proximity of the catheter to the nerves. 25   parameters resulted in a significant reduction in phrenic nerve stimulation (see Supplementary material online, Video S2).
We observed a higher incidence of the event when ablation was performed in the LSPV, which aligns with the location of the phrenic nerve in swine.However, in previous and our research, neither long-term functional damage nor structural damage caused by pulsed electric fields has been verified. 25,37Gross pathology and long-term assessment did not indicate nerve damage, even though the electric field strength exceeded the therapeutic setting when performed directly adjacent to the nerve. 37pathology results also supported this conclusion.It is speculated that the physiological electrical nerve signalling may have been interfered with or intensified by the transient external electrical field, resulting in the overactivation and unanticipated contraction of the diaphragm muscles, with an absence of irreversible damage.
Based on our experimental results, when the catheter was ablated in a spindle shape rather than a lotos shape, phrenic nerve capture occurred more frequently.Possibly, this is due to the smaller electrode spacing in the spindle shape compared with the lotos shape.According to basic research, the delivery energy is negatively correlated with electrode spacing, 24 which may explain the observation.According to this, real-time electrode spacing calculations and current density estimation based on three-dimensional mapping and computational system may be convenient for operators in practical use and represent an important direction for future research.

Histopathology in our study
In histopathology, the lesions were transmural, continuous, homogenous, and endocardial sparing, with the preservation of the structural integrity of the heart.Fibrous collagen and inflammation were involved in the process.No thrombus or injury to the arterial, venular, or nerve structures was observed.These findings are consistent with the previously described characteristics of the non-thermal ablation mechanism of PFA. 24,38

Limitations
In this study, safety and chronic lesions were demonstrated for a relatively short-term follow-up period (up to 30 days), while long-term follow-up studies, such as those conducted for 3 months, are necessary to provide a comprehensive evaluation of durability and safety.
As to histopathology, there were only four longitudinal sections per targeted vein.Segments that were not ablated or did not have transmural lesions were likely missed, resulting in an overestimation of circumferentiality and transmurality.Additionally, it is noteworthy that acute pathology was performed within 30 min after the procedure when the injury foci were not fully developed.Further studies are required to identify acute inflammatory reactions and other acute pathological changes at 7 days post procedure.
For safety, there may be a need for quantitative studies investigating the factors affecting the capture of the phrenic nerve, which are not included in our study and will be investigated in the future.Furthermore, the impact of PFA on ganglionated plexi (GP) and the cardiac autonomic nervous system (ANS) was not investigated in this study.It has been reported that classic PVI procedures using thermal ablations (RF or CB) have an effect on GPs and the ANS. 39][44] Therefore, comparative experiments are necessary to determine whether PFA damages the ANS and whether it impacts the recurrence of AF.
Last but not least, as humans and swine have different anatomical and physiological states, there is a limitation in predicting the efficacy and safety of PFA in humans.Clinical evaluations should be conducted separately and with caution.

Conclusions
Excellent PVI efficacy and acceptable safety can be achieved by this novel lotos PFA catheter and PFA system.It is capable of creating transmural myocardial lesions, while sparing the coronary arteries and the phrenic nerve.Furthermore, it can isolate the PVs without acute or chronic PV stenosis and oesophagus injury.

Figure 1
Figure 1 The study catheter and the pulse parameter used for bipolar PFA.(A1-A3) Multi-electrode catheter gradually deploying into a basic pose (A1), a spindle pose (A2), and a lotos pose (A3).with an 11 Fr shaft.(B) Details of the fully deployed lotos pose (the distal portion is 28 or 31 mm in diameter), shown with 16 electrodes in total.(C and D) Details of a biphasic pulses set.A full cycle consists of a pair of diphasic pulses.PFA, pulsed field ablation.

Figure 2
Figure 2 views.(A and B) PFA catheter deployed over-the-wire in LSPVs of swine in the spindle (A) and the lotos (B) pose through a deflectable sheath.Sensing and pacing catheters were placed in the coronary sinus.(C and D) A PFA catheter deployed over-the-wire in RSPVs of swine in the spindle (C ) and the lotos (D) pose through a deflectable sheath.Sensing and pacing catheters were placed in the coronary sinus.LSPV, left superior pulmonary vein; PFA, pulsed field ablation; RSPV, right superior pulmonary vein.

Table 1 Figure 3
Figure 3 Representative electrograms and electroanatomic maps.(A) PV potential disappearing immediately following ablation.(B and C) Changes of pulmonary vein electrograms obtained from the study catheter before and immediately after PFA in the LSPV (B) and RSPV (C), (D and E), Representative voltage maps performed at baseline, immediately after ablation, and 30-day follow-up showing the extent and chronic lesion of isolation in the LSPV and RSPV.LSPV, left superior pulmonary vein; PFA, pulsed field ablation; RSPV, right superior pulmonary vein.

'Figure 4
Figure 4 Fluoroscopic views and intracardiac echocardiographic views for safety assessment.(A and B) Baseline, immediately post ablation, and 30-day follow-up venous angiography of the LSPV (A) and RSPV (B) showing no venous stenosis.The dotted line denotes pulmonary veins.(C ) A view of the coronary artery before and 8 min after ablation.(D) Intracardiac echocardiographic view of PVs during ablation with spindle pose (D1) and lotos pose (D2), showing no microbubbles.LSPV, left superior pulmonary vein; RSPV, right superior pulmonary vein.

aFigure 5
Figure 5 Gross pathology and histopathology of PFA lesions.(A) A PFA lesion in the LSPV with the contiguous broad lesion, shown in gross appearance prior to formalin fixation.The lesion can be identified as white areas of discolouration and is outlined with the dotted line.(B-D) Representative histopathologic images of the chronic cohort of transmural PFA lesion in the LSPV (B and C) and RSPV (D) with structural framework preserved and endocardial sparing, which was conducted 30 days after the procedure.Haemorrhage and myocardium inflammation can be seen.(C) A higher magnification image of the black box outlined in B, demonstrating a sparing of the arteries, veins (arrowhead), and nerves (arrow).(E and F) Representative histopathologic images of target PV in the acute cohort, demonstrating oedema and mild neoendocardium (arrowhead in F). (E) A close-up of the black box area in (F), showing the preserved structural framework with vessels (black arrowhead) and nerves (arrow) retention.The black asterisk represents unaffected epicardial adipose tissue.Masson trichrome staining was used in (B-F).LSPV, left superior pulmonary vein; PFA, pulsed field ablation; RSPV, right superior pulmonary vein.

Table 2
Measurements of pulmonary vein diameters at different points of time

Table 3
Events of phrenic nerve capture in different poses Values are n/N (%).Comparison is made between PVs that were subjected to ablation in the spindle and flower pose.LSPV, left superior pulmonary vein; RSPV, right superior pulmonary vein.